**2.1 Microbial culture**

116 Biogas

the O2/H2S supplied (v/v), it was low at a ratio of 5.3 and increased up to 68-78% as the

In a biofiltration system, a gas stream is passed through a packed bed on which pollutantdegrading organisms are immobilized as biofilms. Biotrickling filters use the same principle, but an additional liquid phase will flow through the reactor. In both systems, the microorganisms in the biofilms transform the absorbed H2S by metabolic activity into elemental sulfur or sulfate depending on the amount of available oxygen. Oxygen is thus the key parameter that controls the level of oxidation. Sulfur production (Eq. 1) results from the partial oxidation of sulfide instead of complete oxidation to sulfate (Eq. 2) when oxygen

H2S + 0.5O2 → S0 + H2O (1)

As the performance of a biofiltration system depends on the microbial community present in the reactor, the determination of the microorganism and the microbial activity responsible for the behavior of the process is very important. However, there is still a lack of understanding of the structure and dynamics of microbial communities and the physiological role of the main microbial population as well as the correlation between the global performance of the system with the metabolic activities of the microorganisms involved in the process. This knowledge could allow control of the reactor behavior and the design of enhanced processes to eliminate high concentrations of H2S in the gas phase because the performance of the process depends on the robustness of the microbial

Some authors have characterized microbial population diversity present in different gas phase reactors by analysis of biomarkers such as phospholipid fatty acids (Webster et al., 1997), molecular techniques such as fluorescent *in situ* hybridization (FISH) (Moller et al., 1996), cloning and sequencing of ribosomal RNA genes (Roy et al., 2003), terminal restriction fragment length polymorphism (Maestre et al., 2009) and denaturing gradient gel electrophoresis (Borin et al., 2006). There are only a few studies in the literature that focused on determining the microbial diversity of microorganisms capable of removing reduced sulfur compounds in biofilters or gas phase bioreactors using molecular biological approaches. Ding et al., 2006, reported the changes in the microbial diversity of a biofiltertreating methanol and H2S. In this study, the biofilter's initial microbial community had a high diversity, but after the biofiltration system was fed with H2S, the microbial diversity decreased to adapt to the low pH and use H2S as an energy source. Maestre et al., 2010 studied and described the bacterial composition of a lab-scale biotrickling filter (BTF) treating high loads of H2S using 16S rRNA gene clone libraries. The authors reported the diversity, the community structure and the changes in the microbial population on days 42 and 189 of reactor operation. The main changes in microbial diversity were observed at the beginning of the process and again when steady state operation was reached (i.e., neutral pH and at an inlet H2S concentration of 2000 ppmv). At steady state, the major sequences associated with SOB included *Thiothrix* spp., *Thiobacillus* spp., and *Sulfurimonas denitrificans*. Additionally, FISH analysis was used to determine the spatial distribution of sulfuroxidizing bacteria (SOB) along the length of the reactor under pseudo-steady state operation. The aerobic species were found to be predominantly along the system, but some

H2S + 2O2 SO4-2 + 2H+ (2)

is limited, as is shown in Equations 1 and 2 (Kennes and Veiga, 2001).

ratio decreased.

communities (Maestre et al., 2010).

Microbial culture in the biofiltration system was adapted to 10% (v/v) acetic acid and 40 mM Na2SO3.5H2O in mineral media and used for VFA and H2S degradation experiments, respectively. In situ immobilization in the biofilter was facilitated by recirculating the solution containing the microbial culture previously adapted (Ramirez-Saenz et al., 2009).
